A new replication-defective, acute transforming retrovirus (3611-MSV) was recently isolated from mouse and molecularly cloned. Two gag-containing polyproteins (P75 and P90) are found in nonproducer transformed cells. The nucleotide sequence of 1.5 kilobases encompassing the transforming gene (v-raf) of 3611-MSV has been determined. v-raf sequences were found to have been inserted into the p30 region of an ecotropic murine leukemia virus (MuLV), with the concomitant deletion of the 2.4 kilobases extending to the middle of the polymerase gene. A 5-nucleotide direct repeat exists at each end of the v-raf sequences. A single nucleotide deletion, 10 bases upstream from the acquired sequences, places the oncogene in an open reading frame terminated by an amber triplet around 180 nucleotides from the 3 feet onc/MuLV junction. From the deduced amino acid sequence, a hybrid gag-raf polyprotein would have a molecular weight of around 75 kilodaltons. Consistent with the gag-x structure, we find that only the P75 polyprotein is modified by the fatty acid myristate, whereas only the P90 polyprotein is glycosylated. Comparison of the deduced v-raf amino acid sequence with other oncogenes revealed domains homologous to v-src and/or v-mos. c-raf-related sequences have been localized to human chromosome 3, a chromosome whose alterations are associated with small cell lung carcinoma and other familiar carcinomas. Human cells derived from small cell carcinomas express v-raf-specific RNA, as do other established human cell lines. Further characterization of v-raf specific proteins and mRNA will be presented.